Title: Airway Submucosal Gland Function in a Porcine Cystic Fibrosis Model Submucosal glands (SMGs) secrete fluid and mucous into the airways and this function is thought to play an important role in lung microbial clearance and defense. SMGs express abundant antibacterial factors and also appear to serve as a stem cell niche for the proximal airways. Because submucosal glands express abundant CFTR and demonstrate defects in fluid secretion in CF human airways, it has been hypothesized that these structures are important targets for the treatment of CF lung disease. However, the lack of a CF animal model whose SMG distribution and structure are similar to those of humans has hindered definitive resolution of this hypothesis. We propose the use of several innovative in vitro and ex vivo approaches, based on the newly developed CF pig model, to address whether SMG dysfunction in the absence of CFTR indeed hinders airway defenses. Aim 1 will characterize the extent of SMG dysfunction in the CF pig model using single gland assays. These studies will assess the electrolyte composition, pH, viscosity, and production rate of glandular fluid secretions in response to known SMG secretory agonists. These experiments will lay the necessary foundation for extrapolating the data gathered in this new CF model to known SMG functions in humans. Additionally, this aim will test the novel hypothesis that CFTR-dependent glandular secretions alter the activity of surface epithelial chloride channels. Aim 2 seeks to define the contribution of CFTR to antibacterial properties associated with glandular secretions. These studies will use ex vivo vascularized xenograft models of CF and non-CF pig proximal airways with or without SMGs. Additionally, we will generate a novel xenograft model with defined CFTR expression in surface airway epithelial and/or SMG components. These models will be used to test two central hypotheses: 1) SMGs are the primary source of antibacterial factors required for maintaining airway sterility, and 2) CFTR expression in SMGs is necessary for gland-mediated innate immunity in the proximal airway. Aim 3 will determine whether the glandular stem cell niche associated with SMG ducts is altered in the absence of CFTR and/or airway infection. Such studies are highly relevant to understanding potential defects in airway repair and proliferation caused by persistent bacterial infection in the CF lung, as well as the basis of disease induced changes in the composition of SMG cell types that may also influence innate immunity in the airway. Together, these aims will attempt to resolve the question of whether SMGs are necessary targets for the treatment of CF lung disease.